TLR-4 and Sustained Calcium Agonists Synergistically Produce Eicosanoids Independent of Protein Synthesis in RAW264.7 Cells
| Autor: | Daren Stephens, Raymond A. Deems, Edward A. Dennis, Matthew W. Buczynski, Rebecca C. Bowers-Gentry, Andrej Grkovich |
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| Rok vydání: | 2007 |
| Předmět: |
Biochemistry & Molecular Biology
Cell signaling Time Factors 1.1 Normal biological development and functioning Medical and Health Sciences Models Biological Biochemistry Phospholipases A Cell Line Mice chemistry.chemical_compound Adenosine Triphosphate Models Underpinning research Animals Receptor Molecular Biology Phospholipase A Arachidonate 5-Lipoxygenase biology Purinergic receptor NF-kappa B Cell Biology Biological Sciences Biological Lipids Toll-Like Receptor 4 Gene Expression Regulation chemistry Eicosanoid Prostaglandin-Endoperoxide Synthases Chemical Sciences Arachidonate 5-lipoxygenase biology.protein Eicosanoids Calcium Arachidonic acid Eicosanoid Production |
| Zdroj: | The Journal of biological chemistry, vol 282, iss 31 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m701831200 |
| Popis: | Arachidonic acid is released by phospholipase A(2) and converted into hundreds of distinct bioactive mediators by a variety of cyclooxygenases (COX), lipoxygenases (LO), and cytochrome P450s. Because of the size and diversity of the eicosanoid class of signaling molecules produced, a thorough and systematic investigation of these biological processes requires the simultaneous quantitation of a large number of eicosanoids in a single analysis. We have developed a robust liquid chromatography/tandem mass spectrometry method that can identify and quantitate over 60 different eicosanoids in a single analysis, and we applied it to agonist-stimulated RAW264.7 murine macrophages. Fifteen different eicosanoids produced through COX and 5-LO were detected either intracellularly or in the media following stimulation with 16 different agonists of Toll-like receptors (TLR), G protein-coupled receptors, and purinergic receptors. No significant differences in the COX metabolite profiles were detected using the different agonists; however, we determined that only agonists creating a sustained Ca(2+) influx were capable of activating the 5-LO pathway in these cells. Synergy between Ca(2+) and TLR pathways was detected and discovered to be independent of NF-kappaB-induced protein synthesis. This demonstrates that TLR induction of protein synthesis and priming for enhanced phospholipase A(2)-mediated eicosanoid production work through two distinct pathways. |
| Databáze: | OpenAIRE |
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